qseC Mutant Research Articles

Deleting qseC downregulates virulence and promotes cross-protection in Pasteurella multocida

QseC, a histidine sensor kinase of the QseBC two-component system, acts as a global regulator of bacterial stress resistance, biofilm formation, and virulence. The function of QseC in some bacteria is well understood, but not in Pasteurella multocida. We found that deleting qseC in P. multocida serotype A:L3 significantly down-regulated bacterial virulence. The mutant had significantly reduced capsule production but increased resistance to oxidative stress and osmotic pressure. Deleting qseC led to a significant increase in qseB expression. Transcriptome sequencing analysis showed that 1245 genes were regulated by qseC, primarily those genes involved in capsule and LPS biosynthesis and export, biofilm formation, and iron uptake/utilization, as well as several immuno-protection related genes including ompA, ptfA, plpB, vacJ, and sodA. In addition to presenting strong immune protection against P. multocida serotypes A:L1 and A:L3 infection, live ΔqseC also exhibited protection against P. multocida serotype B:L2 and serotype F:L3 infection in a mouse model. The results indicate that QseC regulates capsular production and virulence in P. multocida. Furthermore, the qseC mutant can be used as an attenuated vaccine against P. multocida strains of multiple serotypes.

QseC Mediates Osmotic Stress Resistance and Biofilm Formation in Haemophilus parasuis.

Haemophilus parasuis is known as a commensal organism discovered in the upper respiratory tract of swine where the pathogenic bacteria survive in various adverse environmental stress. QseC, a histidine protein kinase of the two-component regulatory systems CheY/QseC, is involved in the environmental adaptation in bacteria. To investigate the role of QseC in coping with the adverse environment stresses and survive in the host, we constructed a qseC mutant of H. parasuis serovar 13 strain (ΔqseC), MY1902. In this study, we found that QseC was involved in stress tolerance of H. parasuis, by the ΔqseC exhibited a decreased resistance to osmotic pressure, oxidative stress, and heat shock. Moreover, the ΔqseC weakened the ability to take up iron and biofilm formation. We also found that the QseC participate in sensing the epinephrine in environment to regulate the density of H. parasuis.

QseB mediates biofilm formation and invasion in Salmonella enterica serovar Typhi.

QseB is a response regulator of the QseBC two-component system (TCS) which is associated with quorum sensing and functions as a global regulator of flagella, biofilm formation, and virulence. The function of QseB and its interaction with QseC has been the subject of study in some organisms, however, little work was done in Salmonella enterica serovar Typhi (S. Typhi). The objective of this study was to investigate the effect of QseB on biofilm formation and virulence in S. Typhi. It showed that the biofilm formation ability of qseC mutant was limited as compared to the wild type strain. We also show overexpression of qseB was in a qseC mutant. Interestingly, deletion of qseB in a qseC mutant restored a wild type phenotype. These results suggested that QseB may account for the impaired biofilm formation in the absence of QseC. Furthermore, deletion of qseB in wild type cells decreased biofilm formation, whereas overexpression of qseB in wild type cells increased biofilm formation. Quantitative real-time PCR also revealed the up-regulation of some fimbria-associated genes in a qseB overexpression strain. These results indicate that QseB may enhance biofilm formation in the presence of QseC. Taken together, we hypothesize that QseB has dual regulatory functions which are dependent upon its cognate sensor. Additionally, invasion of HeLa cells was enhanced in qseB mutant but attenuated in a qseC mutant compared with wild-type. The β-galactosidase activity of invF::lacZ was increased in qseB mutant but decreased in qseC mutant which was consistent with invasion results. In conclusion, QseB may have dual regulatory functions concerning biofilm formation and plays a negative role in virulence of S. Typhi.

The role of the sensor kinase, QseC, an adrenergic receptor of Escherichia coli, in bacterial translocation during hemorrhagic shock.

Hemorrhagic shock results in ileal mucosa damage and intestinal bacterial translocation. Additionally, during hemorrhagic shock, norepinephrine levels increase. Past research has shown that the QseC sensor kinase of Escherichia coli modulates the quorum-sensing response to epinephrine and norepinephrine. Therefore, the aim of our study was to examine whether the absence of the ability of E. coli to sense epinephrine/norepinephrine would attenuate the bacterial translocation to extraintestinal organs in a rat model of hemorrhagic shock. An E. coli MC1000 qseC mutant was constructed, and E. coli MC1000 and MC1000ΔqseC with streptomycin resistance were used to track bacterial translocation after gavage in rats. A rat model of nonlethal hemorrhagic shock was used. The rats were divided into six groups: controls (SS), rats that received a sham shock and MC1000 (M-SS), rats that received a sham shock and MC1000ΔqseC (Δ-SS), rats that received a hemorrhagic shock alone (HS), rats that received a hemorrhagic shock and MC1000 (M-HS), and rats that received a hemorrhagic shock and MC1000ΔqseC (Δ-HS). We found the incidence of bacterial translocation in the M-HS rats was higher than in the Δ-HS rats. The observed effects seem to be largely dependent on the inability to sense epinephrine/norepinephrine and the decreased motility of E. coli MC1000ΔqseC. Therefore, a role for E. coli sensing epinephrine/norepinephrine in the pathophysiology of bacterial translocation following hemorrhagic shock is proposed. The demonstration of such an effect would suggest a new mechanism for the development of shock-induced sepsis.

The Role of the QseC Quorum-Sensing Sensor Kinase in Epinephrine-Enhanced Motility and Biofilm Formation by Escherichia coli

Biofilms play a pivotal role in infections related to devices. Biofilm formation in Escherichia coli is mediated by the quorum-sensing E. coli regulator C (QseC), the histidine sensor kinase that can sense epinephrine (EPI)/norepinephrine (NE). In this study, we evaluate the role of the QseC quorum-sensing sensor kinase in epinephrine-enhanced motility and biofilm formation by E. coli. An E. coli MC1000 qseC mutant was constructed. We investigated the role of the QseC in the formation of biofilms on the surface of medical-grade polyvinyl chloride using the E. coli K-12 MC1000 strain as well as a corresponding qseC mutant. Addition of EPI/NE increased biofilm formation by wild-type K-12 MC1000 but not by the isogenic qseC mutant. Scanning confocal laser microscopy corroborated these results by showing that EPI/NE addition significantly increased biofilm's thickness. As expected, the addition of EPI/NE to the qseC mutant, which lacks the ability to sense the hormones, failed to stimulate biofilm formation. Since EPI/NE addition increased bacterial motility, we proposed that their stimulatory effects on biofilm formation occur by enhancing bacterial motility and altering biofilm architecture. We also found that EPI/NE regulate motility and the biofilm phenotype via QseC, as motility was diminished and biofilm formation was significantly decreased in a qseC deletion mutant. These results indicate that EPI/NE induce E. coli biofilm formation on the surface of polyvinyl chloride through QseC. Cross-talk between E. coli (quorum sensing) and host hormones may explain the pathogen-caused opportunistic infections that occur in patients with prosthetic devices used during hormone level fluctuations in the host.

The putative sensor kinase QseC of Salmonella enterica serovar Typhi can promote invasion in the presence of glucose

Salmonella enterica serovar Typhi (S. Typhi) causes a human-restricted systemic infection called typhoid fever. It invades intestinal epithelial cells requiring expression of genes located at SPI-1. QseC is a regulator of quorum sensing that initiates transcription of a series of flagellar genes in reponse to AI-3 in Escherichia coli. By sequencing analysis, we have found that the homolog of the putative qseBC (t3098, t3099) in a wild-type strain of S. Typhi shares 98% and 99% identity with that of S. enterica serovar Typhimurium (S. Typhimurium), respectively. Glucose is not only an energy source compound but also an important environmental signaling molecule for enteropathogenic bacteria. In this study, we found that a series of invasion-associated genes in SPI-1 were downregulated in the qseC mutant of S. Tyhpi compared to the wild-type in the presence of glucose, by the genomic microarray assay. Results of qRT-PCR were consistent with the microarray data and suggest that the transcriptional regulation of invasion-associated genes by QseC is glucose-dependent and the regulation is not dependent on QseB. A qseC mutant cultured in the presence of glucose condition was impaired in invasion of epithelial cells. These data indicate that QseC can promote invasion in S. Typhi in the presence of glucose.

The Salmonella enterica serovar Typhimurium QseB response regulator negatively regulates bacterial motility and swine colonization in the absence of the QseC sensor kinase

Salmonella enterica serovar Typhimurium ( S. Typhimurium) responds to the catecholamine, norepinephrine by increasing bacterial growth and enhancing motility. In this study, iron with or without the siderophore, ferrioxamine E also enhanced bacterial motility. Iron-enhanced motility was growth-rate dependent, while norepinephrine-enhanced motility was growth-rate independent. The outer membrane catecholate receptors, IroN, FepA and CirA (required for norepinephrine-enhanced growth) were not required for norepinephrine-enhanced motility, nor was ExbD of the energy-transducing TonB-ExbB-ExbD ferri-siderophore uptake system. Examination of the QseBC two-component system revealed that qseB and qseBC mutants have motility phenotypes similar to wild-type S. Typhimurium, while motility of the qseC mutant was significantly decreased ( P < 0.01 ). Each mutant of the QseBC system, as well as mutants of qseE and pmrA, responded to norepinephrine with increased motility, suggesting that other genes are involved in norepinephrine-enhanced motility of S. Typhimurium. In the swine host, fecal shedding of the qseBC mutant was similar to wild-type S. Typhimurium, whereas fecal shedding of the qseC mutant was significantly decreased ( P < 0.01 ). Our data indicate that, in a qseC mutant, the QseB response regulator decreases motility and swine colonization; inactivation of the qseBC operon restores these bacterial phenotypes, classifying QseB as a negative regulator of bacterial motility and swine colonization.

Francisella novicida Forms In Vitro Biofilms Mediated by an Orphan Response Regulator

Francisella tularensis is associated with water and waterways and infects many species of animals, insects, and protists. The mechanism Francisella utilizes to persist in the environment and in tick vectors is currently unknown. We have demonstrated for the first time that Francisella novicida, a model organism of F. tularensis, forms a biofilm in vitro. Selected F. novicida transposon mutants were tested for their ability to form biofilm compared to the wildtype F. novicida strain. Mutation of the putative qseB gene led to an impairment in the ability to form biofilm with no impairment in bacterial growth. A qseC mutant had impaired growth but demonstrated a marked impairment in biofilm production. Mutation in capC affected both bacterial growth and biofilm formation, but no biofilm production impairment was seen with capB or pilE mutants. A deletion mutant in the orphan response regulator FTN_1465, which we propose is the putative QseB, formed significantly less biofilm than the wildtype. When FTN_1465 was complemented back into the deletion mutant, biofilm formation was restored. Thus, the orphan response regulator FTN_1465 is an important factor in biofilm production in vitro in F. novicida. These results demonstrate that Francisella species are able to form biofilms in vitro, suggesting that biofilm formation may be important for the lifecycle of this organism.

QseC‐mediated dephosphorylation of QseB is required for expression of genes associated with virulence in uropathogenic Escherichia coli

Bacteria sense environmental cues and regulate gene expression accordingly so as to persist in diverse niches. QseC is a membrane sensor kinase shown in enterohemorrhagic Escherichia coli to respond to host and bacterial signals by phosphorylating the QseB response regulator at residue D51, resulting in QseB activation and presumably upregulation of virulence genes. We studied QseBC in uropathogenic E. coli (UPEC). UPEC establish infection by colonizing and invading bladder cells. After invasion, UPEC can escape into the cytoplasm where they can form intracellular bacterial communities. Deletion of qseC significantly attenuated intracellular bacterial community formation and virulence, whereas paradoxically qseB deletion did not impact pathogenesis. We found that QseB upregulates its own expression in the qseC mutant, arguing that it is activated even in the absence of QseC. However, expression of QseB, but not a QseB_D51A mutant, in the absence of QseC resulted in downregulation of type 1 pili, curli and flagella. We observed similar phenotypes with enterohemorrhagic E. coli, showing that this is not a UPEC-specific phenomenon. Target gene expression is restored when QseC is present. We discovered that QseC has phosphatase activity required for QseB dephosphorylation. Thus, the QseC phosphatase capacity is critical for modulating QseB activity and subsequent gene expression.

The role of the QseC quorum-sensing sensor kinase in colonization and norepinephrine-enhanced motility of Salmonella enterica serovar Typhimurium

Transcriptional analysis of Salmonella enterica serovar Typhimurium ( S. Typhimurium) in the presence of the mammalian hormone norepinephrine revealed up-regulation of genes in the flagellar and chemotaxis regulon. Motility assays confirmed enhanced motility of wild-type S. Typhimurium in the presence of norepinephrine that could be blocked by the α-adrenergic antagonist, phentolamine. Furthermore, a mutation in the qseC gene, encoding the sensor kinase of the two-component QseBC quorum-sensing system, also diminished motility of S. Typhimurium. To investigate the role of S. Typhimurium QseC in vivo, 13-week old pigs were intranasally inoculated with equal concentrations (1×10 9 CFU) of wild-type S. Typhimurium and a qseC mutant. Over a 1-week competitive index experiment, the qseC mutant displayed decreased colonization of the gastrointestinal tract compared to the wild-type parent strain. Thus, this study has identified a role for the QseBC quorum-sensing signal transduction system in motility and swine colonization of S. Typhimurium. Cross-talk between cell–cell communication systems in Salmonella (quorum sensing) and host hormones may explain opportunistic behaviors of the pathogen, such as immune evasion and stress-induced recrudescence of Salmonella, during fluctuations of host hormone levels.

Quorum sensing Escherichia coli regulators B and C (QseBC): a novel two-component regulatory system involved in the regulation of flagella and motility by quorum sensing in E. coli.

Quorum sensing is a cell-to-cell signalling mechanism in which bacteria secrete hormone-like compounds called autoinducers. When these auto-inducers reach a certain threshold concentration, they interact with bacterial transcriptional regulators, thereby regulating gene expression. Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 as well as E. coli K-12 produces the autoinducer-2 (AI-2), which is synthesized by the product of the luxS gene, and previous work from our laboratory has shown that genes encoding the EHEC type III secretion system were activated by quorum sensing. Recently, by hybridizing an E. coli K-12 gene array with cDNA synthesized from RNA extracted from EHEC strain 86-24 and its isogenic luxS mutant, we observed that other potential virulence-associated factors, such as genes encoding the expression and assembly of flagella, motility and chemotaxis, were also activated by quorum sensing. The array data also indicated that several genes encoding putative E. coli regulators were controlled by quorum sensing. In this report, we describe a two-component system regulated by quorum sensing that shares homology with Salmonella typhimurium PmrAB, which we have named quorum sensing E. coli regulator B and C (QseBC). The qseBC genes, previously identified only as open reading frames b3025 and b3026, are organized in an operon in the E. coli chromosome, with qseB encoding the response regulator and qseC the sensor kinase. We confirmed the regulation of qseBC by quorum sensing using qseB::lacZ transcriptional fusions and characterized the phenotypes of an isogenic qseC mutation in EHEC. This mutant expressed less flagellin and had reduced motility compared with the wild-type and complemented strains. Transcription of flhD, fliA, motA and fliC::lacZ fusions was decreased in the qseC mutant, suggesting that qseBC is a transcriptional regulator of flagella genes. A qseC mutant was also generated in E. coli K-12 strain MC1000 that showed the same phenotypes as the EHEC mutant, indicating that qseBC regulates flagella and motility by quorum sensing in both EHEC and K-12. QseBC activates transcription of flhDC, which is the master regulator for the flagella and motility genes and, in the absence of flhD, QseBC failed to activate the transcription of fliA. Motility of a luxS, but not of a qseC, mutant can be restored by providing AI-2 exogenously as preconditioned media, suggesting that the qseC mutant is unable to respond to AI-2. However, QseC has no effect on the expression of other quorum sensing-controlled genes such as those encoding for the type III secretion system. These data indicate that QseBC is one component of the quorum-sensing regulatory cascade in both EHEC and K-12 that is involved in the regulation of flagella and motility genes, but that additional regulators in this cascade remain to be characterized.